Nitric oxide (NO) is an important cell signaling molecule that regulates blood pressure in endothelial cells, acts as an important molecule in macrophage cells for immune system defense against pathogens, and is a neurotransmitter in neuronal cells. Pathologically produced excesses of neuronal NO, however, have been correlated with almost all neurodegenerative diseases, including Parkinson's disease, Alzheimer's disease, and Huntington's disease. The objective of this proposal is to diminish this excess neuronal NO by inhibition of the neuronal form of the enzyme that produces it, nitric oxide synthase (NOS). The successes we have had in the design of selective neuronal nitric oxide synthase (nNOS) inhibitors during the last funding period will be the springboard for future design and synthesis efforts, using crystallographic images of our inhibitors bound to NOS and structure-based computer design of new structures. At present two of our inhibitors have been found to be very effective at preventing cerebral palsy phenotype in a rabbit model for cerebral palsy. For this indication, administration by injection is acceptable. The important next step taken will be to enhance the oral bioavailability of these compounds while increasing their potency and selectivity for nNOS over the other two isoforms of NOS, endothelial NOS (eNOS) and inducible NOS (iNOS) to minimize side effects and toxicities. One key observation made in the last funding period was an unexpected binding conformation of some of our selective inhibitors, which opens up the door for more bioavailable inhibitors. Once oral bioavailability improves, then these compounds may become first-in-class treatments for Parkinson's, Alzheimer's, and Huntington's diseases. Basic pharmacokinetic properties of the new molecules will be measured to determine their metabolic stability and ability to enter the brain. Neuroprotective studies in a Parkinson rat model will be conducted. The standard treatment for Parkinson's disease is L-dopa; however, prolonged use of this drug produces abnormal involuntary movements (AIMs) or dyskinesia. Orally bioavailable compounds will be tested for their ability to prevent dyskinesia in both rats and primates treated with L-dopa. Finally, further studies will be undertaken to elucidate the mechanism for how NOS catalyzes the oxidation of L-arginine to citrulline and NO using alternative substrates. PUBLIC HEALTH RELEVANCE: Pathologically produced excesses of neuronal nitric oxide have been correlated with almost every neurodegenerative disease, including Parkinson's disease, Alzheimer's disease, and Huntington's disease. The objective of this proposal is to diminish this excess neuronal nitric oxide by synthesis and evaluation of compounds that inhibit the neuronal form of the enzyme that produces nitric oxide, namely, nitric oxide synthase. When accomplished, these compounds could be first-in-class treatments for these neurodegenerative diseases.